The course of an infection with the mammalian pathogen Pseudomonas aeruginosa is, in part, dependent on the expression of well-characterized virulence factors expressed by all strains of the bacteria, but significant strain-to-strain differences exist in the ability of the organism to cause disease. Genomic islands, regions of DNA contained within the bacterial genome that vary between strains, comprise up to 10% of the P. aeruginosa genome, but remain relatively unexplored. The presence or absence of non- conserved virulence factors within these genomic islands may play a role in modulating pathogenicity from strain to strain. Prior studies have identified several genomic islands from P. aeruginosa clinical isolates that enhance the severity of respiratory infections. In this study, the relationship between genomic islands in P. aeruginosa and pathogenicity will be further explored. This will be accomplished by completing the following two specific aims: In Aim 1, bacterial isolates will be obtained from hospital patients with P. aeruginosa bloodstream infections and tested in a mouse model of bacteremia to identify the most virulent strains. The genome of a particularly hypervirulent P. aeruginosa strain will be sequenced and compared to a reference P. aeruginosa strain to identify novel genomic islands in the clinical isolate. In Aim 2, deletion and complementation of genomic islands from a hypervirulent P. aeruginosa isolate will be performed to identify those genomic islands most associated with virulence. The virulence-associated genomic islands will then be subjected to piecemeal deletions, the effects of which will be examined in a mouse model of bacteremia. These studies will identify the specific ORFs within the genomic islands encoding for virulence factors associated with enhanced pathogenicity. Completion of this study will identify novel P. aeruginosa pathogenicity-associated genomic islands and virulence factors, yielding insight into the mechanisms underlying differences in the courses of P. aeruginosa infections. PUBLIC HEALTH RELEVANCE: Successful completion of this proposal will contribute greatly to the advancement of medical science by identifying novel virulence determinants influencing the infectivity of the major human pathogen, P. aeruginosa. The findings from these studies will also offer new targets for future studies examining the course of P. aeruginosa clinical infection, as well as the development of novel therapeutics against P. aeruginosa.